This invention relates generally to methods and compositions for increasing the solubility of biologically active agents in lipophilic media. More particularly, the invention relates to the use of N,N-dinitramide salts as solubilizing agents for ionizable biologically active agents, particularly biologically active agents that when ionized are positively charged. The invention finds utility in a variety of fields, including pharmaceuticals and drug delivery, medical imaging and diagnostics, agrochemical manufacture, and dietary supplement formulation.
Many hydrophobic compounds are not only poorly soluble or even insoluble in water, but are also insoluble or only slightly soluble in lipophilic media. It is very difficult to manufacture useful products with such compounds because their insolubility limits the ability to chemically transform and/or formulate the compounds in any sort of composition. This is particularly problematic with hydrophobic pharmacologically active agents, which if insoluble in both aqueous and lipophilic media have very low oral bioavailability. Furthermore, for those drugs that are intended to be delivered to the brain, and thus be transported across the xe2x80x9cblood-brain barrierxe2x80x9d as will be described below, an insufficient amount of the drug penetrates the blood brain barrier and enters the central nervous system.
For treating pathologic processes inside the central nervous system (CNS), therapeutic benefit derives from availability of the agent inside the CNS. Undesired effects can result from effects of the agent anywhere in the body of the treated organism. The blood-brain barrier, regulates the exchange of materials between the bloodstream and the CNS, and may present a formidable barrier to drug transport into the CNS, thus affecting the effective dose of an agent or precluding its use by systemic administration altogether. As physical breach of the meninges is known to be medically undesirable, the inability of a pharmacologic agent to penetrate the blood-brain barrier can preclude its use altogether or limit its use to only life threatening situations.
The physiological basis for the blood-brain barrier are the brain capillaries, which are comprised of a lining of endothelial cells (Goldstein et al. (1986) Scientific American 255:74-83 (1986); Pardridge (1986) Endocrin. Rev. 7:314-330). The endothelial cells lining the capillaries of the blood-brain barrier are different from those lining capillaries of other tissues. Specifically, they form complex tight junctions, which prevent passage of molecules and ions between cells. The blood-brain barrier is formed by these high-resistance, tight intercellular junctions along with the endothelial cells themselves. The integrated structure forms a continuous wall against the passive movement of many molecules from the blood to the brain. The capillary endothelial cells forming the blood-brain barrier are also different in that they have few pinocytotic vesicles, which allow somewhat unselective transport across the capillary wall in other tissues. Continuous gaps or channels running through the cells, which would allow unrestrained passage of moieties, are also absent in blood-brain barrier capillary lining.
The structure of the blood-brain barrier may be subdivided into two components: the endothelial or capillary barrier and the ependymalbarrier (Banks et al (1991) Pharm. Res. 8:1345-1350). The nature of the mechanism of the penetration of substances through the blood-brain barrier has not been fully explained, but many regulators of brain function, such as cytokines, transferrin, enkephalins and endorphins, are believed capable of traversing the blood-brain barrier from the blood vessels into the brain (Raeissi et al (1989) J. Pharm. Phy. 41:848-852; Zlokovich et al. (1989) Peptides 10:249-254; and Zlokovich, B. (1990) J. Control. Rel. 13:185-201). However, many substances that can affect the CNS including adenosine, xcex2-endorphin and synthetic analogs of endogenous peptides and some excitatory and inhibitor amino acids and trophic factors, penetrate the blood-brain barrier poorly or not at all (Houghten et al (1980) Proc. Natl. Acad. Sci. USA 77:4588-4591; Levin et al. (1987) Biochem. Biophys. Res. Commun. 147:1226-123; Sakane et al. (1989) Int. J. Pharm. 57:77-83). Currently, drugs with little or no blood-brain barrier penetration can only be administered by direct CNS infusion or by implantation of controlled-release polymers (see, e.g., U.S. Pat. No. 4,883,666 to Sabel et al.). Thus, many potentially potent drugs are not clinically useful inside the CNS because of inability to cross the blood-brain barrier in amounts capable of yielding therapeutic levels in the CNS at below toxic systemic doses.
Many pharmacologic agents exert desirable therapeutic effects inside the CNS at attainable systemic levels, but can be employed with only limited therapeutic scope because of severe side effects to peripheral organs and/or the peripheral nervous system. Thus, a need generally exists to reduce the side effects of drugs directed to the CNS by reducing side effects on peripheral organs and tissues by increasing the action inside the blood-brain barrier.
One approach has been to alter the permeability of the blood-brain barrier itself. For instance, some osmotic agents administered peripherally, as by intravenous or intramuscular injection, result in the breach of the blood-brain barrier. Further, some drugs acting on the CNS can alter the permeability properties of the blood-brain barrier for other substances. Cholinomimetic arecolines, for example, have been reported to induce alterations of drug penetration through the blood-brain barrier (Saija et al. (1990) J. Pharm. Pha. 42:135-138). Other drugs that may be administered to change the blood-brain barrier permeability are disclosed in U.S. Pat. Nos. 5,059,415 and 5,124,146 to Neuwelt. Bradykinin is one specific drug with such effects (see U.S. Pat. No. 5,112,596 to Malfroy-Camine). Another method involves administration of permeabilizer peptides such as A-7 or conformational analogs thereof (see Kozarich et al., International Patent Publication No. WO 92/18529). Tomasz et al. (International Patent Publication No. WO 91/16064) administer parenteral injections of purified cell walls or cell wall fragments of eubacteria such as Streptococcus pneumoniae to open the blood-brain barrier, a relatively invasive method.
U.S. Pat. No. 5,260,210 to Rubin et al. discloses a method of increasing the permeability of the blood-brain barrier by administering an agent to reduce or inhibit intracellular cyclic AMP concentrations or to increase cyclic GMP concentrations. One disadvantage of wholesale alteration of blood-brain barrier permeability is the resulting lack of selectivity. Thus, any method of changing the permeability of the blood-brain barrier itself is compromised by the possible entry of unwanted molecules from which the brain is normally protected by the blood-brain barrier. Such methods are thus impracticable due to unpredictable and uncontrollable consequences.
Another approach has been the modification of drug molecules themselves. The properties of the molecule, such as size and pKa, are important to the drug""s ability to penetrate the blood-brain barrier. For example, macromolecules, including folded proteins, do not pass the blood-brain barrier at all. One way of modifying a molecule so as to render it capable of traversing the blood brain barrier involves isolating the active moiety of a macromolecule, i.e., that portion of the molecule responsible for the biologically desirable result, and using only that active moiety. Because size is one of the factors affecting ability of a molecule to traverse the blood-brain barrier, reduced size is employed to enhance the kinetics of penetration of the blood-brain barrier, and consequently increase the likelihood that the smaller molecule may traverse the blood-brain barrier in therapeutically significant amounts. Other modifications to macromolecules to enhance passage through the blood-brain barrier include glycating proteins to enhance penetration of the blood-brain barrier, or formation of a prodrug. U.S. Pat. No. 5,260,308 to Podusio et al. teaches glycating proteins, while U.S. Pat. No. 4,933,324 to Shashoua and related International Patent Publication No. WO 89/07938 disclose formation of a prodrug formed from a fatty acid carrier and a neuroactive drug unable to pass through the blood-brain barrier on its own. A similar system is disclosed in WO 89/07938.
One variant of the preceding approach involves linking the desired pharmacoactive compound to a peptide that can facilitate crossing the blood-brain barrier by transcytosis, as described in U.S. Pat. No. 6,030,941 to Summerton et al. In transcytosis, polarized endothelial cells of the capillary, having distinct apical and basolateral membranes, effect traversal of the cell forming a barrier such as the blood-brain barrier. A compound transported by transcytosis is first taken through the apical membrane in the inner capillary wall into a transcytotic vesicle. Transcytotic vesicles typically have an internal pH of about 6.0. The vesicle then transfers the compound to the basolateral membrane of the endothelial cell, on the outer capillary wall. The compound is then expelled from the transcytotic vesicle and across the blood-brain barrier.
Yet another approach is the implantation of controlled release polymers that release the active ingredient from a matrix-system directly into the nervous tissue. However, this approach is invasive and requires surgical intervention for implantation directly into the brain or spinal cord (see Sabel et al., U.S. Pat. No. 4,883,666).
To overcome these limitations, another approach has been tried in which drug carrier systems are used including liposomes, erythrocyte ghosts, antibody-conjugates, and monoclonal antibody conjugates. One of the major problems in targeted drug delivery is the rapid opsonization and uptake of such injected drug carriers by the reticuloendothelial system (RES), especially by macrophages in the liver and spleen. This obstacle may be partially overcome for liposomal and other carrier systems by incorporation of xe2x80x9cstealthxe2x80x9d lipids, such as phosphatidylinositol, monosialoganglioside.
The aforementioned approaches are rather limited because they are only effective for specific drugs in specific circumstances. Liposomes are probably the least invasive existing method for pharmacological agent delivery across the blood-brain barrier. However, a number of problems are associated with liposomal delivery. For example, liposomes often exhibit severe stability problems and are therefore only of limited clinical use.
Based on these considerations, a long-felt need is apparent for new carrier systems, particularly systems that are capable of transporting molecules that do not penetrate the blood-brain barrier using liposomal delivery. Methods, preparations and drug delivery systems that permit drugs to traverse are likewise needed. Enhancing penetration of the blood-brain barrier should also, ideally, have reduced or minimal peripheral side effects, while at the same time allowing for full therapeutic effect in the nervous system.
There is a specific need for a method and system for transporting lipophilic cations across the blood-brain barrier, as many CNS active agents and other nervous system agents are amine drugs, the term xe2x80x9caminexe2x80x9d used loosely to refer to compounds containing a tetravalent nitrogen moiety or a nitrogen moiety capable of being protonated. The present invention is addressed to the aforementioned needs in the art, and provides for effective enhancement of an ionizable (e.g., an amine) compound""s lipophilicity and hence solubility in a lipophilic medium, in turn facilitating passage across the blood-brain barrier.
The present method is useful not only in conjunction with delivery of CNS active agents, but also with a host of other pharmacologically active agents, including metal-based drugs.
The invention is not, however, limited to use in drug delivery. The invention finds utility in a host of contexts where there is a need to increase the solubility of a biologically active agent in a lipophilic medium. For example, the invention is useful in conjunction with increasing the lipophilic solubility of medical imaging and diagnostic agents, as well as the lipophilic solubility of xe2x80x9cnon-pharmacologicalxe2x80x9d nutrients such as dietary supplements, herbal extracts, and the like. In addition, in the agricultural field, many agricultural chemicals (or xe2x80x9cagrochemicalsxe2x80x9d), including insecticides, herbicides, fungicides and other pesticides, have little or no aqueous solubility, and are only slightly soluble in lipophilic solvents.
Accordingly, in one embodiment, the invention provides a method for enhancing the solubility of an ionizable compound in a lipophilic medium by admixing the ionizable compound with an effective solubility enhancing amount of an N,N-dinitramide salt. The ionizable compound may be in either ionized or un-ionized form prior to admixture, and if in ionized form, in association with an anionic counterion. The ionizable compound, upon ionization, gives rise to a biologically active cationic species that becomes ionically associated with the N,N-dinitramide anion following admixture. The biologically active cationic species may be a pharmacologically active cation.
Preferred ionizable compounds are ionizable pharmacologically active agents, e.g., those which are centrally acting agents such as CNS amines or other nervous system agents, such as: sympathomimetic amines; neuroprotective and neuroregenerative agents, including neurotrophic factors; neuroactive amino acids and peptides; neurotransmitters; agents to treat neurodegenerative disorders; CNS and respiratory stimulants; and drugs that selectively modify CNS function. The ionizable pharmacologically active agent may also be a metal-based drug or a dietary supplement such as a vitamin, a mineral, or another nutritional supplement.
The ionizable compounds may also be medical imaging or diagnostic agents, or an agricultural chemical agent such as a pesticide.
In a related embodiment, the invention provides novel salts formed by reaction of the ionizable compounds with an N,N-dinitramide salt, wherein the salts comprise the biologically active cation of the ionizable compound is in association with N,N-dinitramide anion. The salts have significantly enhanced solubility in lipophilic media relative to that of the ionizable compound.
In a further embodiment, methods of use are provided. One such method is a process for enhancing the penetration of the blood-brain barrier by a pharmacologically active agent, wherein the method involves coadministering the active agent with an N,N-dinitramide salt or coadministering the active agent as an N,N-dinitramide salt.